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Abstract:

We set up a label-free direct binding assay for the detection of noncoding RNAs. The assay is based on nanomechanical cantilever arrays for the detection of surface stress induced by immobilized biomolecules and their interaction partners. We used various means to significantly reduce the drift of the cantilever readout that was a prominent feature in experiments with readout in stationary fluid before and after sample injection. Major improvements were achieved by focusing on a faster system equilibration (for instance temperature control and diffusion independence). Experimental protocols were improved to provide user-friendly and less time-consuming measurements. Further enhancements were achieved by, for example, using pre-gold-coated cantilever array wafers compared to individually prepared ones and a directly implemented data analysis tool as real-time feature of the measurement software. We have demonstrated picomolar specific biomarker target detection and can easily distinguish modified targets with single-nucleotide mismatches that hybridize with lower affinity.